Structural basis for FEN-1 substrate specificity and PCNA-mediated activation in DNA replication and repair
Structural basis for FEN-1 substrate specificity and PCNA-mediated activation in DNA replication and repair
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DOI：
10.1016/s0092-8674(03)01036-5
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发表时间：
2004-01-09
期刊：
影响因子：
64.5
通讯作者：
Tainer, JA
中科院分区：
文献类型：
--
作者：
Chapados, BR;Hosfield, DJ;Tainer, JA
Flap EndoNuclease-1（FEN-1）和增殖细胞核抗原（PCNA）是DNA复制和修复的核心。为了阐明FEN-1特异性和PCNA激活的分子基础，我们在此报道FEN-1：DNA和PCNA：FEN-1-肽复合物的结构，沿着荧光共振能量转移（FRET）和突变结果。FEN-1结合未配对的3' DNA末端（3'瓣），打开并扭结DNA，并促进柔性螺旋夹的构象闭合以促进5'切割特异性。FEN-1和PCNA中非结构化C-末端区域的排序产生了分子间β折叠界面，其直接连接FEN-1的相邻PCNA和DNA结合区域，并表明PCNA如何刺激FEN-1活性。DNA和蛋白质的构象变化，复合物的复杂结构，FRET，和突变的结果支持酶-PCNA的路线和扭结的DNA支点，出现适合协调旋转的扭结的DNA中间体之间的酶定位的三个PCNA结合位点。
Flap EndoNuclease-1 (FEN-1) and the processivity factor proliferating cell nuclear antigen (PCNA) are central to DNA replication and repair. To clarify the molecular basis of FEN-1 specificity and PCNA activation, we report here structures of FEN-1:DNA and PCNA: FEN-1-peptide complexes, along with fluorescence resonance energy transfer (FRET) and mutational results. FEN-1 binds the unpaired 3' DNA end (3' flap), opens and kinks the DNA, and promotes conformational closing of a flexible helical clamp to facilitate 5' cleavage specificity. Ordering of unstructured C-terminal regions in FEN-1 and PCNA creates an intermolecular beta sheet interface that directly links adjacent PCNA and DNA binding regions of FEN-1 and suggests how PCNA stimulates FEN-1 activity. The DNA and protein conformational changes, composite complex structures, FRET, and mutational results support enzyme-PCNA alignments and a kinked DNA pivot point that appear suitable to coordinate rotary handoffs of kinked DNA intermediates among enzymes localized by the three PCNA binding sites.